Diversidad de ninfas de Leptohyphidae (Ephemeroptera) en el río Quenane-Quenanito, piedemonte llanero colombiano / Diversity of Leptohyphidae (Ephemeroptera) nymphs in the Quenane-Quenanito river, Colombian plain foothills

Resumen Introducción: Varias presiones antrópicas sufren los ecosistemas acuáticos del piedemonte llanero en Colombia. La respuesta a estresores ambientales aún se desconoce en organismos bioindicadores como Leptohyphidae. Objetivo: Determinar la diversidad de ninfas de Leptohyphidae del río Quenane-Quenanito, en dos periodos hidrológicos contrastantes y su relación con algunas variables fisicoquímicas. Métodos: En diciembre (2014) y febrero (2015) se recolectaron organismos con red Surber en seis estaciones a lo largo del río. Se analizó la diversidad alfa y beta y se aplicó análisis de redundancia y modelos lineales generalizados con el fin de establecer la relación entre los taxones y las variables ambientales. Resultados: Se identificaron 369 organismos pertenecientes a cuatro géneros (Amanahyphes, Traverhyphes, Tricorythopsis y Tricorythodes), dos especies y ocho morfoespecies. Se reporta por primera vez para el departamento del Meta Amanahyphes saguassu. Se registró la mayor diversidad de ninfas en la transición a la sequía y la mayor abundancia en sequía. La diversidad beta señaló que la configuración del ensamblaje cambia a nivel espacial y temporal. Conclusiones: Los organismos de Leptohyphidae prefieren hábitats de corrientes, particularmente en el periodo de sequía, donde hallan alimento (hojarasca, detritos) y refugio para establecerse exitosamente; actividades antrópicas como la urbanización afectan notablemente la diversidad. La alta diversidad registrada en este pequeño río de piedemonte llanero refleja la necesidad de incrementar este tipo de trabajos y esfuerzos de recolección de material de estudio en la región.

Abstract Introduction: Various anthropic pressures affect the aquatic ecosystems of the foothills of Colombia. The response to environmental stressors is still unknown in bioindicator organisms such as Leptohyphidae. Objective: To determine the diversity of Leptohyphidae nymphs of the Quenane-Quenanito river, in two contrasting hydrological periods and its relationship with some physicochemical variables. Methods: In December (2014) and February (2015), organisms were collected with a Surber net at six stations along the current. Alpha and beta diversity was analyzed and redundancy analysis and generalized linear model were applied to establish the relationship between taxa and environmental variables. Results: Were identified 369 organisms belonging to four genera (Amanahyphes, Traverhyphes, Tricorythopsis, and Tricorythodes), two species, and eight morphospecies. Amanahyphes saguassu is reported for the first time for the Meta department. High diversity of Leptohyphidae nymphs was recorded in the transition to drought season and greater abundance in drought. Beta diversity indicated that the configuration of the assemblage changes spatially and temporally. Conclusions: Leptohyphidae organisms prefer fast habitats, particularly in the dry period where they find food (leaf litter, detritus) and shelter to establish themselves successfully; anthropic activities such as urbanization notably affect diversity. The high diversity recorded in this small river in the foothills of the plains reflects the need to increase this type of works and collection efforts of study material in the region.

Predictability of ecological and evolutionary dynamics in a changing world.

Ecological and evolutionary predictions are being increasingly employed to inform decision-makers confronted with intensifying pressures on biodiversity. For these efforts to effectively guide conservation actions, knowing the limit of predictability is pivotal. In this study, we provide realistic expectations for the enterprise of predicting changes in ecological and evolutionary observations through time. We begin with an intuitive explanation of predictability (the extent to which predictions are possible) employing an easy-to-use metric, predictive power PP(t). To illustrate the challenge of forecasting, we then show that among insects, birds, fishes and mammals, (i) 50% of the populations are predictable at most 1 year in advance and (ii) the median 1-year-ahead predictive power corresponds to a prediction R 2 of only 20%. Predictability is not an immutable property of ecological systems. For example, different harvesting strategies can impact the predictability of exploited populations to varying degrees. Moreover, incorporating explanatory variables, accounting for time trends and considering multivariate time series can enhance predictability. To effectively address the challenge of biodiversity loss, researchers and practitioners must be aware of the information within the available data that can be used for prediction and explore efficient ways to leverage this knowledge for environmental stewardship.

Coevolution between eggs and sperm of insects.

Sexual selection is known to play a major role in the evolution of insect sperm size, whereas natural selection is thought to be a major driver of insect egg size. Despite these differing forms of selection operating, it is possible coevolution between male and female gametes can occur owing to their vital interactions during fertilization. We tested egg-sperm coevolution in insects and found that longer sperm correlated to longer and wider eggs. Moreover, the size of the entry point of sperm into insect eggs (micropyles), was positively related to the diameter of sperm, on average being approximately three times the diameter of the sperm. This suggests a function in reducing and channelling sperm entry, but potentially still leaving space for movement. Our work suggests that greater attention needs to be paid to egg-sperm interactions prior to the point of fertilization as they may influence the evolution of gametes.

Overview and Evolution of Insect Fibroin Heavy Chain (FibH).

The FibH gene, crucial for silk spinning in insects, encodes a protein that significantly influences silk fiber mechanics. Due to its large size and repetitive sequences, limited known sequences of insect FibH impede comprehensive understanding. Here, we analyzed 114 complete FibH gene sequences from Lepidoptera (71 moths, 24 butterflies) and 13 Trichoptera, revealing single-copy FibH in most species, with 2-3 copies in Hesperinae and Heteropterinae (subfamily of skippers). All FibH genes are structured with two exons and one intron (39-45 bp), with the second exon being notably longer. Moths exhibit higher GC content in FibH compared to butterflies and Trichoptera. The FibH composition varies among species, with moths and butterflies favoring Ala, Gly, Ser, Pro, Gln, and Asn, while Trichoptera FibH is enriched in Gly, Ser, and Arg, and has less Ala. Unique to Trichoptera FibH are Tyr, Val, Arg, and Trp, whereas Lepidoptera FibH is marked by polyAla (polyalanine), polySer (polyserine), and the hexapeptide GAGSGA. A phylogenetic analysis suggests that Lepidoptera FibH evolved from Trichoptera, with skipper FibH evolving from Papilionoidea. This study substantially expands the FibH repertoire, providing a foundation for the development of artificial silk.

HOFE: an interactive forensic entomological database.

The significance of entomological evidence in inferring the time, location and cause of death has been demonstrated both theoretically and practically. With the advancement of sequencing technologies, reports have emerged on necrophagous insects' nuclear genomes, transcriptomes, proteomes and mitochondrial genomes. However, within the field of forensic entomology, there is currently no available database that can integrate, store and share the resources of necrophagous insects. The absence of a database poses an inconvenience to the application of entomological evidence in judicial practice and hampers the development of the forensic entomology discipline. Given this, we have developed the Home Of Forensic Entomology database, encompassing 10 core functional modules: Home, Browse, Mitochondria, Proteome, JBrowse, Search, BLAST, Tools, Case base and Maps. Notably, the 'Tools' module enables multiple sequence alignment analysis (Muscle), homologous protein prediction (Genewise), primer design (Primer), large-scale genomic analysis (Lastz), Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analysis, as well as expression profiling (PCA Analysis, Hcluster and Correlation Heatmap). In addition, the present database also works as an interactive platform for researchers by sharing forensic entomological case reports and uploading data and material. This database provides potential visitors with a comprehensive function for multi-omics data analysis, offers substantial references to researchers and criminal scene investigators and facilitates the utilization of entomological evidence in court. Database URL: http://ihofe.com/.

Biochemical characterisation and production kinetics of high molecular-weight (HMW) putative antibacterial proteins of insect pathogenic Brevibacillus laterosporus isolates.

BACKGROUND: Bacterial genomes often encode structures similar to phage capsids (encapsulins) and phage tails which can be induced spontaneously or using genotoxic compounds such as mitomycin C. These high molecular-weight (HMW) putative antibacterial proteins (ABPs) are used against the competitive strains under natural environment. Previously, it was unknown whether these HMW putative ABPs originating from the insect pathogenic Gram-positive, spore-forming bacterium Brevibacillus laterosporus (Bl) isolates (1821L, 1951) are spontaneously induced during the growth and pose a detrimental effect on their own survival. Furthermore, no prior work has been undertaken to determine their biochemical characteristics. RESULTS: Using a soft agar overlay method with polyethylene glycol precipitation, a narrow spectrum of bioactivity was found from the precipitated lysate of Bl 1951. Electron micrographs of mitomycin C- induced filtrates showed structures similar to phage capsids and contractile tails. Bioactivity assays of cell free supernatants (CFS) extracted during the growth of Bl 1821L and Bl 1951 suggested spontaneous induction of these HMW putative ABPs with an autocidal activity. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis of spontaneously induced putative ABPs showed appearance of ~ 30 kDa and ~ 48 kDa bands of varying intensity across all the time intervals during the bacterial growth except in the initial hours. Statistically, spontaneously induced HMW putative ABPs of Bl 1951 exhibited a significant decrease in the number of viable cells of its producer strain after 18 h of growth in liquid. In addition, a significant change in pH and prominent bioactivity of the CFS of this particular time period was noted. Biochemically, the filtered supernatant derived from either Bl 1821L or Bl 1951 maintained bioactivity over a wide range of pH and temperature. CONCLUSION: This study reports the spontaneous induction of HMW putative ABPs (bacteriocins) of Bl 1821L and Bl 1951 isolates during the course of growth with potential autocidal activity which is critically important during production as a potential biopesticide. A narrow spectrum of putative antibacterial activity of Bl 1951 precipitate was found. The stability of HMW putative ABPs of Bl 1821L and Bl 1951 over a wide range of pH and temperature can be useful in expanding the potential of this useful bacterium beyond the insecticidal value.

Expansion and Storage of Insect Cells and Virus Stocks.

Healthy insect cell cultures are critical for any method described in this book, including making productive baculovirus banks, protein or AAV expression, and determining viral titers. This chapter describes cell maintenance in shake flasks using serum-free conditions and the expansion of virus stocks from a single plaque purified virus. Insect cells can be passaged over multiple generations, but as the cells may undergo changes over multiple passages, limiting the use of your cells to a defined number of passages such as 50 passages is recommendable. Baculovirus stocks once created using serum-free media are not very stable at 4-8 °C. This chapter also includes a simple method to store cells from an early cell passage and your virus stock in liquid nitrogen.

A Time and Cost-Effective Pipeline for Expression Screening and Protein Production in Insect Cells Based on the HR-Bac Toolbox to Generate Recombinant Baculoviruses.

The baculovirus expression vector system (BEVS) is recognized as a powerful platform for producing challenging proteins and multiprotein complexes both in academia and industry. Since a baculovirus was first used to produce heterologous human IFN-ß protein in insect cells, the BEVS has continuously been developed and its applications expanded. We have recently established a multigene expression toolbox (HR-bac) composed of a set of engineered bacmids expressing a fluorescent marker to monitor virus propagation and a library of transfer vectors. Unlike platforms that rely on Tn7-medidated transposition for the construction of baculoviruses, HR-bac relies on homologous recombination, which allows to evaluate expression constructs in 2 weeks and is thus perfectly adapted to parallel expression screening. In this chapter, we detail our standard operating procedures for the preparation of the reagents, the construction and evaluation of baculoviruses, and the optimization of protein production for both intracellularly expressed and secreted proteins.

Construction of Recombinant Baculoviruses.

The success of using the insect cell-baculovirus expression technology (BEST) relies on the efficient construction of recombinant baculovirus with genetic stability and high productivity, ideally within a short time period. Generation of recombinant baculoviruses requires the transfection of insect cells, harvesting of recombinant baculovirus pools, isolation of plaques, and the expansion of baculovirus stocks for their use for recombinant protein production. Moreover, many options exist for selecting the genetic elements to be present in the recombinant baculovirus. This chapter describes the most commonly used homologous recombination systems for the production of recombinant baculoviruses, as well as strategies to maximize generation efficiency and recombinant protein or baculovirus production. The key steps for generating baculovirus stocks and troubleshooting strategies are described.

RNAi-Mediated Silencing in the Insect Cell-Baculovirus Expression System.

RNA interference (RNAi) serves as an indispensable tool for gene function studies and has been substantiated through extensive research for its practical applications in the baculovirus expression vector system (BEVS). This chapter expands the RNAi toolkit in insect cell culture by including small interfering RNA (siRNA) in the protocol, in addition to the conventional use of double-stranded RNA (dsRNA). This chapter also brings attention to key design and reporting considerations, based on Minimum Information About an RNAi Experiment (MIARE) guidelines. Recommendations regarding online tools for dsRNA and siRNA design are provided, along with guidance on choosing suitable methods for measuring silencing outcomes.

Adaptive Laboratory Evolution to Improve Recombinant Protein Production Using Insect Cells.

Adaptive laboratory evolution (ALE) is a powerful tool for enhancing the fitness of cell lines in specific applications, including recombinant protein production. Through adaptation to nonstandard culture conditions, cells can develop specific traits that make them high producers. Despite being widely used for microorganisms and, to lesser extent, for mammalian cells, ALE has been poorly leveraged for insect cells. Here, we describe a method for adapting insect High Five and Sf9 cells to nonstandard culture conditions via an ALE approach. Aiming to demonstrate the potential of ALE to improve productivity of insect cells, two case studies are demonstrated. In the first, we adapted insect High Five cells from their standard pH (6.2) to neutral pH (7.0); this adaptation allowed to improve production of influenza virus-like particles (VLPs) by threefold, using the transient baculovirus expression vector system. In the second, we adapted insect Sf9 cells from their standard culture temperature (27 °C) to hypothermic growth (22 °C); this adaptation allowed to improve production of influenza VLPs by sixfold, using stable cell lines. These examples demonstrate the potential of ALE for enhancing productivity within distinct insect cell hosts and expression systems by manipulating different culture conditions.

Production of Recombinant Viral Antigens Using the Baculovirus-Insect Cell Expression System.

Insect cell expression has been successfully used for the production of viral antigens as part of commercial vaccine development. As expression host, insect cells offer advantage over bacterial system by presenting the ability of performing post-translational modifications (PTMs) such as glycosylation and phosphorylation thus preserving the native functionality of the proteins especially for viral antigens. Insect cells have limitation in exactly mimicking some proteins which require complex glycosylation pattern. The recent advancement in insect cell engineering strategies could overcome this limitation to some extent. Moreover, cost efficiency, timelines, safety, and process adoptability make insect cells a preferred platform for production of subunit antigens for human and animal vaccines. In this chapter, we describe the method for producing the SARS-CoV2 spike ectodomain subunit antigen for human vaccine development and the virus like particle (VLP), based on capsid protein of porcine circovirus virus 2 (PCV2d) antigen for animal vaccine development using two different insect cell lines, SF9 & Hi5, respectively. This methodology demonstrates the flexibility and broad applicability of insect cell as expression host.

Purification Process for a Secreted Protein Produced in Insect Cells.

The Baculovirus Expression Vector System (BEVS) is used with cultured insect cells to produce a wide variety of heterologous proteins, which can be secreted into the culture medium during the transient infection process (Smith et al. Mol Cell Biol 12:2156-2165, 1983). When the infection process is complete, centrifugation is often used to separate the desired protein from the spent insect cells. The desired product in the harvested supernatant is contaminated with baculovirus, amino acids, lipids, detergents, oils, lysed cells from the infection process, genomic DNA from the insect cells, and proteases due to the lytic nature of the baculovirus infection process and many other contaminants (Ikonomou et al. Appl Microbiol Biotechnol 62:1-20, 2003). All these contaminants that are present in the centrifuged supernatant with the desired secreted protein make the initial chromatographic capture step critical for effective purification of the desired protein. A purification scheme will be outlined for a slightly acidic secreted protein using cation exchange chromatography (Lundanes et al. Chromatography: basic principles, sample preparations and related methods, 1st edn. Wiley, 2013).

Nonviral Platform for Expression of Recombinant Protein in Insect Cells.

Nonviral transfection has been used to express various recombinant proteins, therapeutics, and virus-like particles (VLP) in mammalian and insect cells. Virus-free methods for protein expression require fewer steps for obtaining protein expression by eliminating virus amplification and measuring the infectivity of the virus. The nonviral method uses a nonlytic plasmid to transfect the gene of interest into the insect cells instead of using baculovirus, a lytic system. In this chapter, we describe one of the transfection methods, which uses polyethyleneimine (PEI) as a DNA delivery material into the insect cells to express the recombinant protein in both adherent and suspension cells.

The contribution of commonly consumed edible insects to nutrition security in the Eastern D.R. Congo.

Edible insects are perceived as an incredible opportunity to mitigate the major challenge of sustainably producing healthy foods for a growing world population in the face of climate change uncertainties over the coming decade. In this study, we assessed the nutrient composition and sensory properties of Acheta domesticus, Apis mellifera, Gnathocera trivittata, Gryllotalpa africana, Imbrasia epimethea, Imbrasia oyemensis, Locusta migratoria, Macrotermes subhylanus, Nomadacris septemfasciata, Rhyncophorus phoenicis, Ruspolia differens and Rhynchophorus ferrugineus consumed in Eastern D. R. Congo. The investigated edible insects are highly appreciated and nutritious, with proteins (20.67-43.93 g/100 g) and fats (14.53-36.02 g/100 g) being the major macro-nutrients, proving their potential to improve diets through food enrichment. The high potassium (24-386.67 mg/100 g), sodium (152-257.82 mg/100 g), magnesium (32-64 mg/100 g), iron (5.3-16.13 mg/100 g), calcium (25-156.67 mg/100 g) and zinc (11-19.67 mg/100 g) content make the assessed edible insects a useful mineral-containing ingredient for preventing undernutrition in countries which are plagued by micronutrient deficiencies. A scatter plot of matrices and Pearson's correlations between sensory attributes and nutritional composition showed a negative correlation (r = - 0.45) between protein and appearance. While no strong correlation was observed between nutritional attributes and sensory acceptance, a positive correlation was observed between potassium and aroma (r = 0.50), after-taste (r = 0.50) and acceptability (r = 0.52). Principal component analysis results indicated that the two axes accounted for up to 97.4% of the observed variability in the nutrient composition and sensory attributes of commonly consumed edible insects in the Eastern D. R. Congo. Given the significant delicacy and nutritional potential of edible insects highlighted in this paper, households can rely on the latter to meet their nutritional needs rather than conventional livestock, thus contributing to environmental and financial security through local business opportunities.

Rat poison's long reach.

Supertoxic rodenticides are building up inside unintended targets, including birds, mammals, and insects. Scientists want to understand the damage-and limit it.

The Collapsible Light Trap: a portable Pennsylvania Light Trap for collecting aquatic insects.

A Collapsible Light Trap (CLT) for collecting insects, particularly aquatic insects, is described here. CLT is a modified Pennsylvania Light Trap with the advantage of being collapsible and lightweight to be carried in a small backpack and very easy to set up in the field. CLT is equipped with LED light strip wrapped around a PVC tube and can be connected to a regular 12 V / 7 Ah battery, running for more than 48 uninterrupted hours. Complete CLT weighs 0.8-1.0 kg, depending on the metal used, and the battery weighs around 2 kg, being easily transportable to more remote collecting areas. Over the years, CLTs have been used for collecting and describing the diversity of aquatic insects from Brazil, particularly caddisflies. Depending on the locality, only one trap for one night can collect over a thousand insect specimens and more than 200 individuals of caddisflies.